Human cancer cells typically contain somatically altered genomes, characterized by mutation, amplification, or deletion of critical genes. In addition, the DNA template from human cancer cells often displays somatic changes in DNA methylation. See, e.g., E. R. Fearon, et al, Cell 61:759 (1990); P. A. Jones, et al., Cancer Res. 46:461 (1986); R. Holliday, Science 238:163 (1987); A. De Bustros, et al., Proc. Natl. Acad. Sci. USA 85:5693 (1988); P. A. Jones, et al., Adv. Cancer Res. 54:1 (1990); S. B. Baylin, et al., Cancer Cells 3:383 (1991); M. Makos, et al., Proc. Natl. Acad. Sci. USA 89:1929 (1992); N. Ohtani-Fujita, et al., Oncogene 8:1063 (1993).
DNA methylases transfer methyl groups from the universal methyl donor S-adenosyl methionine to specific sites on the DNA. Several biological functions have been attributed to the methylated bases in DNA. The most established biological function is the protection of the DNA from digestion by cognate restriction enzymes. This restriction modification phenomenon has, so far, been observed only in bacteria.
Mammalian cells, however, possess different methylases that exclusively methylate cytosine residues on the DNA that are 5′ neighbors of guanine (CpG). This methylation has been shown by several lines of evidence to play a role in gene activity, cell differentiation, tumorigenesis, X-chromosome inactivation, genomic imprinting and other major biological processes (Razin, A., H., and Riggs, R. D. eds. in DNA Methylation Biochemistry and Biological Significance, Springer-Verlag, N.Y., 1984).
In eukaryotic cells, methylation of cytosine residues that are immediately 5′ to a guanosine, occurs predominantly in CpG poor loci (Bird, A., Nature 321:209 (1986)). In contrast, discrete regions of CG dinucleotides called CpG islands (CGi) remain unmethylated in normal cells, except during X-chromosome inactivation and parental specific imprinting (Li, et al., Nature 366:362 (1993)) where methylation of 5′ regulatory regions can lead to transcriptional repression. For example, de novo methylation of the Rb gene has been demonstrated in a small fraction of retinoblastomas (Sakai, et al., Am. J. Hum. Genet., 48:880 (1991)), and a more detailed analysis of the VHL gene showed aberrant methylation in a subset of sporadic renal cell carcinomas (Herman, et al., Proc. Natl. Acad. Sci. U.S.A., 91:9700 (1994)). Expression of a tumor suppressor gene can also be abolished by de novo DNA methylation of a normally unmethylated 5′ CpG island. See, e.g., Issa, et al., Nature Genet. 7:536 (1994); Merlo, et al., Nature Med. 1:686 (1995); Herman, et al., Cancer Res., 56:722 (1996); Graff, et al., Cancer Res., 55:5195 (1995); Herman, et al., Cancer Res. 55:4525 (1995).
Identification of the earliest genetic and epigenetic changes in tumorigenesis is a major focus in molecular cancer research. Diagnostic approaches based on identification of these changes can allow implementation of early detection strategies, tumor staging and novel therapeutic approaches targeting these early changes, leading to more effective cancer treatment. The present invention addresses these and other problems.